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目的探讨低密度脂蛋白(LDL)对大鼠近端肾小管上皮细胞(NRK-52E)转分化的影响及其作用机制。方法将体外培养的NRK-52E细胞分为4组:正常对照组、洛伐他汀(Lov,1μmol/L)组、LDL(250 mg/L)刺激组、LDL(250mg/L)+Lov(1μmol/L)组。应用倒置显微镜观察细胞形态学改变;RT-PCR检测NRK-52E细胞α-平滑肌肌动蛋白(α-SMA)、E-钙粘蛋白(E-cadherin)、转化生长因子β1(TGF-β1)、Smad2、Smad3和整合素链接酶(ILK)mRNA表达;West-ern blot检测α-SMA、E-cadherin及磷酸化Smad2/3(p-Smad2/3)蛋白表达;ELISA检测上清液TGF-β1。结果加入LDL培养24 h后,NRK-52E细胞由立方形或多角形变为长梭形,α-SMA、TGF-β1、Smad2、Smad3和ILK mRNA的表达上调,E-cadherin mRNA的表达下调,与对照组比较差异均有统计学意义(P<0.01);α-SMA和p-Smad2/3蛋白表达量显著高于对照组(P<0.01),E-cadherin蛋白表达量显著低于对照组(P<0.01);细胞上清液TGF-β1浓度显著高于对照组(P<0.01)。LDL+Lov组与LDL组比较,NRK-52E细胞α-SMA、TGF-β1、Smad2、Smad3、ILK mRNA的表达及α-SMA、p-Smad2/3的蛋白表达均明显降低,E-cadherin mRNA和蛋白表达明显增强。结论LDL可诱导肾小管上皮细胞转分化,其机制可能是激活TGF-β1/Smads/ILK信号通路,而Lov能部分阻断LDL诱导的小管上皮细胞转分化。
Objective To investigate the effect of low density lipoprotein (LDL) on transdifferentiation of proximal tubular epithelial cells (NRK-52E) in rats and its mechanism. Methods NRK-52E cells cultured in vitro were divided into 4 groups: normal control group, LOV (1μmol / L) group, LDL (250mg / L) / L) group. The morphological changes of cells were observed by inverted microscope. The expressions of α-SMA, E-cadherin and TGF-β1 in NRK-52E cells were detected by RT- The expressions of Smad2, Smad3 and ILK mRNA were detected by Western blotting. The expressions of α-SMA, E-cadherin and p-Smad2 / 3 protein were detected by Western-ern blot. The expressions of TGF- . RESULTS: After cultured with LDL for 24 h, the expression of α-SMA, TGF-β1, Smad2, Smad3 and ILK mRNA was up-regulated and the expression of E-cadherin mRNA was down-regulated in NRK-52E cells from cuboid or polygonal to long fusiform (P <0.01). The expression of α-SMA and p-Smad2 / 3 protein in the control group was significantly higher than that in the control group (P <0.01), and the expression of E-cadherin protein in the control group was significantly lower than that in the control group P <0.01). The concentration of TGF-β1 in the cell supernatant was significantly higher than that in the control group (P <0.01). Compared with LDL group, the expressions of α-SMA, TGF-β1, Smad2, Smad3 and ILK mRNA and the expressions of α-SMA and p-Smad2 / 3 in NRK-52E cells were significantly decreased in LDL + Lov group And protein expression was significantly enhanced. Conclusion LDL can induce renal tubular epithelial cell transdifferentiation. Its mechanism may be to activate TGF-β1 / Smads / ILK signaling pathway, while Lov can partially block LDL-induced tubular epithelial cell transdifferentiation.